Canadine (tetrahydroberberine) as a GyrB-Targeting antibacterial lead against Acinetobacter baumannii: Integrated phenotypic, computational, and enzymatic evidence.

The global rise of antimicrobial resistance among Gram-negative hospital pathogens urgently demands new antibacterial candidates supported by both phenotypic activity and mechanistic evidence. Here, canadine (tetrahydroberberine), a principal alkaloid of Hydrastis canadensis L., was evaluated as an antibacterial lead against berberine. Target nomination via reverse pharmacophore mapping (PharmMapper), followed by bacterial-target filtering, prioritized DNA gyrase subunit B (GyrB) for mechanistic investigation. Whole-cell antibacterial activity, assessed by broth microdilution against a panel of pathogenic bacteria, revealed that canadine exhibited consistently superior activity to berberine, including enhanced inhibition of Acinetobacter baumannii. Structure-based docking against the A. baumannii GyrB crystal structure (PDB: 7PQL), validated by redocking of the co-crystallized inhibitor (RMSD = 1.68 Å), yielded a favorable docking score for canadine (-10.5 kcal/mol), approaching that of the co-crystallized ligand (-10.8 kcal/mol) and outperforming berberine. Molecular dynamics simulations (100 ns) demonstrated stable canadine-GyrB complexes with lower conformational deviation and reduced energetic fluctuation relative to berberine. Interaction-energy decomposition identified van der Waals forces as the primary stabilizing contributions, with electrostatic interactions serving as secondary modulators. Binding free-energy calculations further supported favorable canadine-GyrB association (ΔG = -9.57 kcal/mol). In vitro E. coli DNA gyrase inhibition assays confirmed greater inhibitory potency for canadine over berberine. Collectively, these findings establish canadine as a mechanistically credible GyrB-targeting antibacterial lead with superior performance to berberine across phenotypic, computational, and biochemical assessments, warranting further optimization through structure-activity relationship studies, selectivity profiling, and translational evaluation against multidrug-resistant A. baumannii.